We have used 18 neutralizing monoclonal antibodies (MAbs) specific for the respiratory syncytial virus (RSV) fusion (F) glycoprotein to construct operational, topologic, and functional epitope maps of the F glycoprotein. The MAbs define 16 neutralization epitopes which are organized into three non-overlapping antigenic sites (A, B, and C) and one bridge site (AB). Neutralization and fusion- inhibition tests of the MAbs indicated that antigenic sites A, AB, and C correspond to functional domains of the F protein involved in syncytium-formation, whereas antigenic site B does not participate directly in the fusion process. This conclusion is reinforced by two additional observations. First, MAb-resistant variants selected with site B MAbs produce normal plaques, whereas those selected with MAbs binding epitopes in sites A and B produce pinpoint plaques. Second, epitopes in antigenic sites, A, AB, and C (which correspond to functional domains) were less variable among RSV clinical isolates than were epitopes in antigenic site B. Sequence analysis of the F genes of antibody-resistant mutants will allow us to identify amino acid residues which are responsible for binding neutralizing antibodies and which participate in syncytium- formation.